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NeuroMolecular Medicine

, Volume 19, Issue 4, pp 525–540 | Cite as

Mutation in GNE Downregulates Peroxiredoxin IV Altering ER Redox Homeostasis

  • Pratibha Chanana
  • Gayatri Padhy
  • Kalpana Bhargava
  • Ranjana AryaEmail author
Original Paper

Abstract

GNE myopathy is a rare neuromuscular genetic disorder characterized by early adult onset and muscle weakness due to mutation in sialic acid biosynthetic enzyme, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE). More than 180 different GNE mutations are known all over the world with unclear pathomechanism. Although hyposialylation of glycoproteins is speculated to be the major cause, but cellular mechanism leading to loss of muscle mass has not yet been deciphered. Besides sialic acid biosynthesis, GNE affects other cellular functions such as cell adhesion and apoptosis. In order to understand the effect of mutant GNE protein on cellular functions, differential proteome profile of HEK293 cells overexpressing pathologically relevant recombinant mutant GNE protein (D207V and V603L) was analyzed. These cells, along with vector control and wild-type GNE-overexpressing cells, were subjected to two-dimensional gel electrophoresis coupled with mass spectrometry (MALDI-TOF/TOF MS/MS). In the study, 10 differentially expressed proteins were identified. Progenesis same spots software revealed downregulation of peroxiredoxin IV (PrdxIV), an ER-resident H2O2 sensor that regulates neurogenesis. Significant reduction in mRNA and protein levels of PrdxIV was observed in GNE mutant cell lines compared with vector control. However, neither total reactive oxygen species was altered nor H2O2 accumulation was observed in GNE mutant cell lines. Interestingly, ER redox state was significantly affected due to reduced normal GNE enzyme activity. Our study indicates that downregulation of PrdxIV affects ER redox state that may contribute to misfolding and aggregation of proteins in GNE myopathy.

Keywords

GNE myopathy UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine kinase Peroxiredoxin IV Sialic acid ROS MERO-GFP ER Stress 

Abbreviations

GNE

UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine kinase

ROS

Reactive Oxygen Species

PrdxIV

Peroxiredoxin-4

PrdxIII

Peroxiredoxin-3

PDI

Protein Disulfide Isomerase

2D-GE

Two-dimensional gel electrophoresis

Ero1

Endoplasmin reticulum oxidoreductase

H2O2

Hyderogen peroxide

MERO-GFP

Mammalian endoplasmic reticulum-localized RedOx-sensitive Green Fluorescent Protein

Notes

Acknowledgements

We acknowledge Prof Fumihiko Urano, Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO, USA, for MERO-GFP construct. We also thank Prof. Alok Bhattacharya (School of Life Sciences, Jawaharlal Nehru University, New Delhi) and Prof. Sudha Bhattacharya (School of Environmental Sciences, Jawaharlal Nehru University, New Delhi) for thoughtful discussions and progressive comments during the project. We thank Mr. Ashok Kumar Sahu, Technical Officer, Advanced Instrumentation Research Facility, JNU for confocal facility.

Funding

This work was supported by the Department of Science and Technology-PURSE-II (DST/SR/PURSE Phase II/11) and University Grants Commission-UPOE II (Project ID: 16), Govt. of India.

Compliance with Ethical Standards

Conflict of interest

Authors declare no conflict interest.

Supplementary material

12017_2017_8467_MOESM1_ESM.docx (880 kb)
Supplementary material 1 (DOCX 880 kb)

References

  1. Abbasi, A., Corpeleijn, E., et al. (2012). Peroxiredoxin 4, a novel circulating biomarker for oxidative stress and the risk of incident cardiovascular disease and all-cause mortality. Journal of the American Heart Association, 1(5), e002956.CrossRefPubMedPubMedCentralGoogle Scholar
  2. Amsili, S., Zer, H., et al. (2008). UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) binds to alpha-actinin 1: Novel pathways in skeletal muscle? PLoS ONE, 3(6), e2477.CrossRefPubMedPubMedCentralGoogle Scholar
  3. Baik, S. C., Kim, K. M., et al. (2004). Proteomic analysis of the sarcosine-insoluble outer membrane fraction of Helicobacter pylori strain 26695. Journal of Bacteriology, 186(4), 949–955.CrossRefPubMedPubMedCentralGoogle Scholar
  4. Broccolini, A., & Mirabella, M. (2015). Hereditary inclusion-body myopathies. Biochimica et Biophysica Acta, 1852(4), 644–650.CrossRefPubMedGoogle Scholar
  5. Choi, M. H., Ow, J. R., et al. (2016). Oxidative stress-mediated skeletal muscle degeneration: Molecules, mechanisms, and therapies. Oxidative Medicine and Cellular Longevity, 2016, 6842568.PubMedGoogle Scholar
  6. Fischer, C., Kleinschnitz, K., et al. (2013). Cell stress molecules in the skeletal muscle of GNE myopathy. BMC Neurology, 13, 24.CrossRefPubMedPubMedCentralGoogle Scholar
  7. Galeano, B., Klootwijk, R., et al. (2007). Mutation in the key enzyme of sialic acid biosynthesis causes severe glomerular proteinuria and is rescued by N-acetylmannosamine. Journal of Clinical Investigation, 117(6), 1585–1594.CrossRefPubMedPubMedCentralGoogle Scholar
  8. Grover, S., & Arya, R. (2014). Role of UDP-N-acetylglucosamine2-epimerase/N-acetylmannosamine kinase (GNE) in beta1-integrin-mediated cell adhesion. Molecular Neurobiology, 50(2), 257–273.CrossRefPubMedGoogle Scholar
  9. Harazi, A., Becker-Cohen, M., et al. (2017). The interaction of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) and alpha-actinin 2 Is altered in GNE myopathy M743T mutant. Molecular Neurobiology, 54(4), 2928–2938.CrossRefPubMedGoogle Scholar
  10. Hinderlich, S., Salama, I., et al. (2004). The homozygous M712T mutation of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase results in reduced enzyme activities but not in altered overall cellular sialylation in hereditary inclusion body myopathy. FEBS Letters, 566(1–3), 105–109.CrossRefPubMedGoogle Scholar
  11. Kanekura, K., Ishigaki, S., et al. (2013). Establishment of a system for monitoring endoplasmic reticulum redox state in mammalian cells. Laboratory Investigation, 93(11), 1254–1258.CrossRefPubMedPubMedCentralGoogle Scholar
  12. Kwofie, M. A., & Skowronski, J. (2008). Specific recognition of Rac2 and Cdc42 by DOCK2 and DOCK9 guanine nucleotide exchange factors. Journal of Biological Chemistry, 283(6), 3088–3096.CrossRefPubMedGoogle Scholar
  13. Li, H., Chen, Q., et al. (2013). Unfolded protein response and activated degradative pathways regulation in GNE myopathy. PLoS ONE, 8(3), e58116.CrossRefPubMedPubMedCentralGoogle Scholar
  14. Ling, L. U., Tan, K. B., et al. (2011). The role of reactive oxygen species and autophagy in safingol-induced cell death. Cell Death and Disease, 2, e129.CrossRefPubMedPubMedCentralGoogle Scholar
  15. Merksamer, P. I., Trusina, A., et al. (2008). Real-time redox measurements during endoplasmic reticulum stress reveal interlinked protein folding functions. Cell, 135(5), 933–947.CrossRefPubMedPubMedCentralGoogle Scholar
  16. Milman Krentsis, I., Sela, I., et al. (2011). GNE is involved in the early development of skeletal and cardiac muscle. PLoS ONE, 6(6), e21389.CrossRefPubMedPubMedCentralGoogle Scholar
  17. Oka, O. B., & Bulleid, N. J. (2013). Forming disulfides in the endoplasmic reticulum. Biochimica et Biophysica Acta, 1833(11), 2425–2429.CrossRefPubMedGoogle Scholar
  18. Padhy, G., Sethy, N. K., et al. (2013). Abundance of plasma antioxidant proteins confers tolerance to acute hypobaric hypoxia exposure. High Altitude Medicine & Biology, 14(3), 289–297.CrossRefGoogle Scholar
  19. Poynton, R. A., & Hampton, M. B. (2014). Peroxiredoxins as biomarkers of oxidative stress. Biochimica et Biophysica Acta, 1840(2), 906–912.CrossRefPubMedGoogle Scholar
  20. Roussel, B. D., Kruppa, A. J., et al. (2013). Endoplasmic reticulum dysfunction in neurological disease. Lancet Neurology, 12(1), 105–118.CrossRefPubMedGoogle Scholar
  21. Schwarzkopf, M., Knobeloch, K. P., et al. (2002). Sialylation is essential for early development in mice. Proceedings of the National Academy of Sciences, 99(8), 5267–5270.CrossRefGoogle Scholar
  22. Sela, I., Milman Krentsis, I., et al. (2011). The proteomic profile of hereditary inclusion body myopathy. PLoS ONE, 6(1), e16334.CrossRefPubMedPubMedCentralGoogle Scholar
  23. Singh, R., & Arya, R. (2016). GNE myopathy and cell apoptosis: A comparative mutation analysis. Molecular Neurobiology, 53(5), 3088–3101.CrossRefPubMedGoogle Scholar
  24. Tateyama, M., Takeda, A., et al. (2003). Oxidative stress and predominant Abeta 42(43) deposition in myopathies with rimmed vacuoles. Acta Neuropathologica, 105(6), 581–585.PubMedGoogle Scholar
  25. Tavender, T. J., & Bulleid, N. J. (2010). Peroxiredoxin IV protects cells from oxidative stress by removing H2O2 produced during disulphide formation. Journal of Cell Science, 123(Pt 15), 2672–2679.CrossRefPubMedPubMedCentralGoogle Scholar
  26. Tavender, T. J., Sheppard, A. M., et al. (2008). Peroxiredoxin IV is an endoplasmic reticulum-localized enzyme forming oligomeric complexes in human cells. Biochemical Journal, 411(1), 191–199.CrossRefPubMedPubMedCentralGoogle Scholar
  27. Trachootham, D., Lu, W., et al. (2008). Redox regulation of cell survival. Antioxidants & Redox Signaling, 10(8), 1343–1374.CrossRefGoogle Scholar
  28. Tsuruta, Y., Furuta, A., et al. (2002). Increased expression of manganese superoxide dismutase is associated with that of nitrotyrosine in myopathies with rimmed vacuoles. Acta Neuropathologica, 103(1), 59–65.CrossRefPubMedGoogle Scholar
  29. Valko, M., Leibfritz, D., et al. (2007). Free radicals and antioxidants in normal physiological functions and human disease. International Journal of Biochemistry & Cell Biology, 39(1), 44–84.CrossRefGoogle Scholar
  30. Varki, A. (2008). Sialic acids in human health and disease. Trends in Molecular Medicine, 14(8), 351–360.CrossRefPubMedPubMedCentralGoogle Scholar
  31. Varki, N. M., & Varki, A. (2007). Diversity in cell surface sialic acid presentations: Implications for biology and disease. Laboratory Investigation, 87(9), 851–857.CrossRefPubMedGoogle Scholar
  32. Wang, Z., Sun, Z., et al. (2006). Roles for UDP-GlcNAc 2-epimerase/ManNAc 6-kinase outside of sialic acid biosynthesis: Modulation of sialyltransferase and BiP expression, GM3 and GD3 biosynthesis, proliferation, and apoptosis, and ERK1/2 phosphorylation. Journal of Biological Chemistry, 281(37), 27016–27028.CrossRefPubMedGoogle Scholar
  33. Weidemann, W., Stelzl, U., et al. (2006). The collapsin response mediator protein 1 (CRMP-1) and the promyelocytic leukemia zinc finger protein (PLZF) bind to UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE), the key enzyme of sialic acid biosynthesis. FEBS Letters, 580(28–29), 6649–6654.CrossRefPubMedGoogle Scholar
  34. Yan, Y., Wladyka, C., et al. (2015). Prdx4 is a compartment-specific H2O2 sensor that regulates neurogenesis by controlling surface expression of GDE2. Nature Communications, 6, 7006.CrossRefPubMedPubMedCentralGoogle Scholar
  35. Zito, E., Melo, E. P., et al. (2010). Oxidative protein folding by an endoplasmic reticulum-localized peroxiredoxin. Molecular Cell, 40(5), 787–797.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  • Pratibha Chanana
    • 1
  • Gayatri Padhy
    • 2
  • Kalpana Bhargava
    • 2
  • Ranjana Arya
    • 1
    Email author
  1. 1.School of BiotechnologyJawaharlal Nehru UniversityNew DelhiIndia
  2. 2.Peptide and Proteomics DivisionDIPAS, DRDODelhiIndia

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